Action potentials regulate the function of the heart by determining beat rate, spread of excitability, and contractility. The action potential in turn is generated by a population of ion channels which undergo conformation changes to alter the permeability of the cell. The ion channels are tuned to provide a certain permeability by the transmembrane voltage, their inherent time dependence, and by intracellular and extracellular modifiers of gating. Life threatening arrhythmias arise in heart tissue through irregularities in the action potential and intercellular conduction. To better understand the origin of arrhythmias and the mechanisms of antiarrhythmic agents it is necessary to understand in detail the ionic conduction mechanisms that underlie the action potential. Five potassium selective channels in chick heart have been identified: 1) a 4pS channel underlying the inward rectifier current most active at hyperpolarized voltages; 2) a 15pS delayed rectifier current channel regulating plateau duration and repolarization; 3) a calcium-activated K channel; 4) a rare 62pS channel active at depolarized voltages; and 5) a muscarinic-gated inward rectifier current in atria. The current proposal seeks funding for further characterization of three of the above channels, specifically with regard to the intracellular modification of gating. Patch clamp methodologies will be used to further characterize the 15pS delayed rectifier channel, the calcium-activated K channel, and the muscarinic-gated inward rectifier. Intracellular regulation of the delayed rectifier will be explored using Beta-adrenergic activation in cell-attached and whole-cell recording. Direct activation by appropriate second messengers will be attempted with inside-out patches. The link between the muscarinic-gated channel of chick atria and the muscarinic receptor will be studied using specific antibodies to G proteins as well as by reconstitution of G proteins in cell-free inside-out patches. The details of the calcium and voltage sensitivity of IK.Ca will be quantified and hormone stimulated and second messenger increases of intracellular calcium explored. By using all the various patch clamp configurations to study the intracellular regulation of K channels in heart, we hope to gain a better understanding of the dynamic control of the action potential.